Adjuvant is a key component in vaccine development. In current avian vaccines, there are some disadvantages existing in the conventional adjuvant such as aluminum hydroxide gel adjuvant and oil adjuvant. For example, the above two adjuvants are chemical adjuvants that cannot enhance specific Th1 cell immune response. Especially, the conventional adjuvant cannot stimulate enough innate and adaptive immune responses against infections.
Therefore, Krieg A. M. et al. found that unmethylated dinucleotide CpG motifs in bacterial deoxyribonucleic acid (DNA) has advantages on stimulating several immune cells to secrete cytokines for enhancements of innate and adaptive immunity. See Krieg A. M. et al., Nature 374:546-549, 1995. The effectiveness of the DNA containing CpG motifs has been confirmed in 1998. The DNA containing CpG motifs has been used as a vaccine adjuvant in 2003. Moreover, the oligodeoxynucleotides (ODN) containing CpG motif can enhance the activities of lymphocytes and antigen-presenting cells (APC), trigger dendritic cell (DC) maturation and antigen-presenting function, and drive immune systems toward Th1 cell immune response against specific antigens, in several large domestic animals such as cattle, pigs, sheep etc. For example, one of the present inventors has provided an ODN containing porcine-specific phosphorothioate (PTO)-modified CpG motif, as well as an immunostimulatory plasmid containing various sets of CpG motifs specifically to pigs.
In general, the mechanisms involved in enhancing immune responses by the ODNs containing CpG motifs (CpG ODN) can be as follows. At first, CpG ODN enhances DC activation, maturation and antigen-presenting function. Secondly, CpG ODN increases DC migration. Thirdly, CpG ODN significantly elevates expression of DC cell markers such as MHC-II, CD40, CD80, CD86, and IL-12 in mice and human. Fourthly, CpG ODN induces priming DC more responsive to antigen-specific Th1 cells. Fifthly, CpG ODN increases CD8□ T cell cytotoxicity activity (CTL) to specific viruses or tumor cells. With application of CpG ODN, CpG ODN itself induces innate immune response to confer protective immune response against infection by viruses, bacteria and extracellular parasites, and B cells are activated by CpG ODN and vaccines to produce antibodies, to activate APC for secreting cytokines such as interferon-γ (IFN-γ), so as to enhance the immune response to vaccines.
However, the prior ODN containing PTO-modified CpG motif DNA synthesized by chemical processes is time-consuming, costs expensively and cannot be produced in mass. The CpG ODN is suitable for an adjuvant instead of PTO-modified CpG ODN. Typically, the CpG ODN contains unmethylated CpG motif, but without replacement of a phosphor atom of a phosphodiester bond in the PTO-modified CpG ODN with a sulfur atom, by which the phosphothioate reduces the degradation rate of the PTO-modified CpG ODN digested by deoxyribonuclease (DNase). In addition, the sequence of CpG ODN is species-specific; the CpG structures between different species are different in immunostimulatory activity. Currently, the sequence of most effective immunoregulatory CpG motif is different between human and mice, and thus specific CpG motifs effective in various species are necessarily confirmed by experiments. The CpG ODN was applied to enhance poultry immunity since 2002. Related researches on the effectiveness of CpG ODN in poultry are mostly involved in evaluation of chemically synthesized CpG ODN in vitro, or focused on antibodies-related humoral immune response in vivo; investigation on the adjuvanicity of CpG ODN on the cell-mediated immune response effectively against viral infection in avians is very limited.
Hence, it is necessary to provide an effective avian immunostimulatory CpG ODN as the DNA adjuvant in avian vaccines, thereby overcoming the disadvantages of the prior DNA adjuvant modified by chemical processes complicatedly.